Project Summary: Antisense oligonucleotide platform for rare genetic diseases of the nervous system Rare diseases affect nearly one in ten people in the United States. Approximately two million of those individuals suffer from rare diseases of the nervous system such as ALS, Fragile X Syndrome, Rett Syndrome, and severe epileptic encephalopathies. Orphan disease variants are often not of interest to large pharmaceutical companies. As a result, hundreds of thousands of patients are left not knowing the cause, potential treatments, or prognosis for their disease creating a large unmet medical need globally. Antisense oligonucleotides or ASOs have several characteristics that make them amenable to a precision medicine approach for treating underlying genetic defects. The FDA has recently approved multiple ASO products for the treatment of rare genetic disease in severe cases unresponsive to traditional drug therapies. These include nusinersen for the treatment of spinal muscular atrophy, eteplirsen for the treatment of Duchene muscular dystrophy and mipomersen for the treatment of homozygous familial hypercholesterolemia. Therapeutic ASOs are generally 15-30 nucleotides in length and complementary to mRNA or pre-spliced mRNA to either inhibit translation through RNAseH mediated mRNA decay or induce exon skipping or inclusion during mRNA splicing. The type of disease-causing variant, whether recessive, dominant gain-of- function or dominant loss-of- function (haploinsufficiency) and intended mode of action of the ASO dictate the design requirement for ASO treatment. Despite the recent clinical successes, ASO design is not rational and requires hundreds of ASO candidates to be synthesized and then evaluated in cell-based assays to identify top ASO candidates. Many of these ASOs often contain undesirable characteristics reducing the probability of positive outcomes in in vivo models and in safety evaluation. In Phase 1, we propose to construct a variant classification algorithm to determine which variants may be amenable to ASO treatment and construct an algorithm to design an appropriate ASO therapy for the associated rare disease variant. We will use ASOs designed through these algorithms to test their efficacies in patient derived iPS cell-derived neurons with causative variants for early-infantile epileptic encephalopathy, a devastating neurological disease associated with rare variants in KCNQ2 and KCNT1, often resulting in significant intellectual development impairment and mortality in childhood. We will expand our ASO platform to new genetic diseases and ASO methodologies in Phase 2 with the goal of developing treatments for patients with rare neurological disorders.